Insect cell lines are used as a culture system for the production of diagnostics and vaccines used in human and veterinary medicine. Many recombinant proteins have been expressed in insect cells that are immunogenically, antigenically, and functionally similar to the native proteins. The desired product is an expressed protein that is produced in large amounts and that is as similar to the natural protein as possible, including necessary post-translation processing and modification. Among the post-translational processing steps that have been shown to occur in insect cells are fatty acid acylation, phosphorylation, and glycosylation (Luckow, V. A. 1995. In: Baculoviruses Expression Systems and Biopesticides, Shuler et al., Eds. Wiley-Liss, New York, N. Y., pages 51–90). Most of the proteins recovered from insect cell cultures, however, migrated faster on SDS-PAGE gels than the native protein, indicating a lower molecular weight because of incomplete post-translational modification.
Baculoviruses as expression vectors are used to produce large amounts of recombinant proteins of medical, pharmaceutical, and veterinary importance in various insect cell lines. Differences in yields of expressed gene products from engineered baculoviruses among cell lines have been reported. Hink et al. (1991) Biotechnol Prog. 7:9–14, compared the expression of three recombinant proteins in twenty-three different cell lines. For each protein, the yield varied among the cell lines and no single cell line produced the highest yields for all three proteins.
In order to facilitate downstream recovery and purification of the expressed recombinant proteins and to reduce costs, insects cells are grown in suspension in large volumes and in medium free of fetal bovine serum (FBS) (Vaughn, J. 1999), In: Encyclopedia of Bioprocess Technology: Fermentation, Biocatalysis, and Bioseparation, Flickinger et al., Eds. John Wiley & Sons, Inc., NY, pages 1444–1457). There are several protein-free media available from commercial suppliers and the yields of recombinant gene products from cells grown in these media generally have been comparable to yields in FBS-supplemented media.
Yields of protein from baculovirus expression vectors in insect cell cultures are reported to be many times higher than those from mammalian cells. The baculovirus expression vector system (BEVS) has been used with hundreds of genes, the majority of which have expressed biologically active proteins (Vaughn, J., supra). The majority of the host cell lines, however, express the recombinant gene products intracellularly. There exists a need in the art for new cell lines that express recombinant gene products extracellularly, and to produce high yields of functional proteins with the desired post-translation modification and proper protein folding.
The invention, as disclosed and described herein, overcomes the shortcomings of the prior art recombinant protein production through the generation of novel insect cell lines that express post translationally modified recombinant proteins extracellularly. The use of the novel cell lines of the invention results in the enhanced expression of expressed recombinant proteins that demonstrate enhanced antigenicity and immunogenicity.